Efforts have been continued to identify and characterize the constituents responsible for the biological activities of the etiologic agent of Rocky Mountain spotted fever, Rickettsia rickettsii. Mice immunized with a soluble rickettsial fraction, purified by affinity chromatography and containing principally the 120-kd surface protein but also detectable amounts of several other components, produced two kinds of antibodies: one specific for RP 120 and one specific for the LPS-like antigen. Since antibodies to the LPS-like antigen do not protect mice, it seems probable that RP 120 was responsible for the immunity developed in mice. RP 120 was found to be hydrophobic in a detergent phase separation experiment; the hydrophobic properties of this protein may account for our difficulties in purifying RP 120. Guinea pigs inoculated with monoclonal antibodies specific for RP 120 or Rp 155 were partially protected against challenge with R. rickettsii. Antibodies to the LPS-like antigen did not protect. Four kinds of monoclonal antibodies were derived from mice inoculated with rickettsiae inactivated at 56 degrees C for 15 min. Three kinds were identical to those obtained from mice given viable rickettsiae; the newly-observed kind recognized heat-resistant (HR) epitopes on RP 155. Antibodies to the HR epitopes did not protect mice from lethal challenge with R. rickettsii. Epitopes recognized by our monoclonal antibodies are distributed widely among spotted fever group rickettsiae; these antibodies could be used to identify all of the species of the group, as well as to differentiate strains within at least one species. In addition, one of the monoclonal antibodies to a HR epitope of RP 155 reacted strongly with the corresponding protein of R. canada, a member of the typhus group. Monoclonal antibodies to the LPS-like antigen reacted with all members of the spotted fever group by ELISA. This antigen probably is the group antigen shared by all members of the group.